The present invention relates to peptide growth factors and their purification from biological tissue. More particularly, the invention is directed to the purification and characterization of low molecular weight (&lt;10,000) peptide growth factors derived from estrogen responsive tissue.
The Government may have rights in this invention pursuant to a funding agreement sponsored by a National Cancer Institute Grant, No. RO1-CA-26617, and an American Cancer Society Grant, No. BC-255.
Polypeptide growth factors have been isolated from organs of various animal species, noncellular plasma fractions, cellular elements of the blood, and cells growing in culture; these activities have been implicated in mammalian cell growth in vivo and in vitro. Several of these mitogens have been purified to homogeneity by procedures that utilize extraction into acidic pH solutions and/or treatment at high temperatures to remove major impurities. The mitogenic species isolated by these methods show molecular weights ranging from 6,000 to 63,000 and primary structures consisting of either one polypeptide chain or two polypeptide chains covalently attached by disulfide bonds.
Major members of the acid-stable family of growth factors are:
1. EGF, epidermal growth factor; PA1 2. MSA, multiplication stimulating activity; PA1 3. IGF I and IGF II, insulin-like growth factors I and II, respectively; PA1 4. SmC, somatomedin C; PA1 5. PDGF, platelet-derived growth factor; PA1 6. TGF, transforming growth factor; and PA1 7. SGF, Sarcoma growth factor.
While, to some degree, members of this group share common properties (i.e. insulin-like responses of IGF I and IGF II), most are distinct molecular entities which, in some cases, interact with different specific cell-surface receptors and in other cases interact with a common receptor.
The role of estrogen inducible growth factors in estrogen responsive growth of mammary tumor cells was first proposed circa 1978. At that time, estrogens were proposed to induce some target tissues to synthesize and/or secrete polypeptide growth factors to enter the circulation and promote the growth of distant mammary target tissue tumors. Potential tissue sources for such factors have been identified as uterus, kidney and pituitary. Investigators designated these growth factors "estromedins" to indicate their role as mediators of the estrogenic growth effects in vivo.
Previously, estrogen responsive tumor cell growth factor activity has been identified in extracts of uteri and kidneys obtained from estrogen-treated rats. Initially, applicants conducted studies to determine whether these organs might be the possible sources of mammary/pituitary cell growth factor activities found in plasma and serum, and to determine whether these activities have a role in the estrogen-responsive growth of mammary and pituitary tumors in vivo. Preliminary data showed that MTW9/PL rat mammary and GH3/C14 rat pituitary tumor cells grew (as measured by cell number increase) in serum free medium supplemented with only u/ml quantities of crude extracts of rat uteri or kidneys.
Since these studies were conducted with heterogeneous preparations, several important questions remained to be resolved. First, the question of whether these growth effects were due to the combination of several relatively non-specific growth and/or survival promoters was unresolved. It seemed possible that more than one mitogenic activity might have been present in the crude extracts, and that together these were responsible for our observed mitogenic effects. Second, the initial observations did not resolve whether the growth factor(s) extracted from the kidney was the same as that extracted from the uterus. Our studies suggested molecular differences since extracts of rat uteri contained an approximately 20 times higher specific activity than identical preparations of rat kidney did, although such differences could have also been related to total growth factor content and not to specific molecular properties.
Applicants in their copending application, Ser. No. 597,980, filed Apr. 6, 1984 have previously described purified peptide growth derived from estrogen responsive uterine and pituitary tissue. The present invention provides separate but related purified peptide growth factors derived from estrogen responsive kidney tissue.